Influenza Vaccination Results in Differential Hemagglutinin Stalk-Specific Fc-Mediated Functions in Individuals Living With or Without HIV

Abstract

Influenza virus hemagglutinin (HA) stalk-specific antibodies have been shown to potently induce Fc-mediated effector functions which are important in protection from disease. In placebo-controlled maternal influenza (MatFlu) vaccination trials of pregnant women living with or without HIV, reduced risk of influenza illness was associated with high HA stalk antibody titers following trivalent inactivated vaccination (TIV). However, the mechanisms of immunity conferred by the HA stalk antibodies were not well understood. Here, we investigated HA stalk-specific Fc effector functions including antibody-dependent cellular phagocytosis (ADCP), antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent complement deposition (ADCD), and FcγRIIa and FcγRIIIa binding in response to seasonal influenza vaccination. These were measured pre- and 1-month post-vaccination in 141 HIV-uninfected women (67 TIV and 74 placebo recipients) and 119 women living with HIV (WLWH; 66 TIV and 53 placebo recipients). In contrast to HIV-uninfected women, where HA stalk-specific ADCP and FcγRIIa binding were significantly boosted, WLWH showed no increase in response to vaccination. HA stalk-specific ADCC potential and FcγRIIIa binding were not boosted regardless of HIV status but were higher in WLWH compared with HIV-uninfected women prior to vaccination. HA stalk-specific ADCD was significantly increased by vaccination in all women, but was significantly lower in the WLWH both pre- and post- vaccination. Co-ordination between HA stalk-specific ADCP and ADCD in WLWH was improved by vaccination. Fc polyfunctionality was enhanced by vaccination in HIV-uninfected women and driven by the HA stalk antibody titers. However, in the WLWH, higher pre-vaccination Fc polyfunctionality was maintained post-vaccination but was decoupled from titer. Overall, we showed differential regulation of Fc effector HA stalk responses, suggesting that HIV infection results in unique humoral immunity in response to influenza vaccination, with relevance for future strategies that aim to target the HA stalk in this population.

Keywords: Fc effector functions; HIV co-infection; antibody- dependent cellular cytotoxicity (ADCC); antibody-dependent cellular phagocytosis (ADCP); antibody-dependent complement deposition (ADCD); hemagglutinin stalk antibodies; influenza vaccination.

Figure 1

A/H1N1 HAI and H1 stalk antibody responses amongst vaccinated women. (A) Pre-vaccination and 1-month post-vaccination hemagglutination inhibition (HAI) titers against A/H1N1 and (B) H1 stalk titers by ELISA, with HIV-uninfected participants (n=67), shown in blue and participants living with HIV (n=66), shown in red. The lines represent the median. Wilcoxon matched-pairs signed rank test used to compare pre- and post-vaccination titers. Mann Whitney U test used to compare responses between vaccine groups. Significant associations shown as ****p < 0.0001; ***p < 0.001; ns,not significant.

Figure 2

Differential boosting of HA stalk-specific Fc-mediated antibody functions amongst vaccinated women. Pre-vaccination and 1-month post-vaccination H1 stalk-specific (A) antibody-dependent cellular phagocytosis (ADCP), (B) antibody-dependent cellular cytotoxicity (ADCC) or FcγRIIIa activation and (C) antibody-dependent complement deposition (ADCD) are shown where HIV-uninfected participants (n=67), shown in blue and participants living with HIV (n=66), shown in red.(D) The fold increases for each of the functions and the proportion (%) of high responders that exceed a 2-fold increase in Fc activity are shown in orange. Low responders, who show reduced enhancement that does not reach a 2-fold increase are shown in black, whilst those not boosted (non-responders) are shown in grey. The lines represent the median. Wilcoxon matched-pairs signed rank tests were used to compare pre-vaccination and post-vaccination functional scores within groups. Mann Whitney U test was used to compare functional scores between the vaccine groups. Fischer’s exact tests were used to compare the proportions of high responders. Significant associations are shown as ****p < 0.0001; ***p < 0.001; **p < 0.01; ns,not significant.

Figure 3

Co-ordination between HAI titers, HA stalk antibodies and HA stalk-specific Fc-mediated functions in response to vaccination. Spearman correlations of Fc effector function and titers in (A) HIV-uninfected pre-vaccination, (B) HIV-uninfected post-vaccination, (C) WLWH pre-vaccination and (D) WLWH post-vaccination groups. The asterisks indicate the following statistical significance, ****p < 0.0001; ***p < 0.001; **p < 0.01; *p < 0.05; ns,not significant. Significant correlations are displayed in red. Non-significant correlations are in grey. The size of the circle is proportional to the Spearman correlation coefficients -0.3 been the smallest and 1 been the largest, indicated inside of circles for correlations that were significant.

Figure 4

HA stalk-specific Fc polyfunctionality scores of Fc-mediated functions amongst vaccinated women. (A) Fc polyfunctionality (determined by the addition of individual standardized Fc function scores), prior to and following vaccination. Dots above 0 indicate Fc polyfunctional individuals, while those below indicate poor Fc polyfunctionality below the mean. The proportion (%) of individuals with Fc polyfunctionality is indicated on the plot. Spearman´s correlation between the Fc polyfunctionality score and HA stalk response in (B) HIV-uninfected women and (C) WLWH. Wilcoxon matched-pairs signed rank test was used to compare pre-vaccination and post-vaccination polyfunctionality scores within groups. Mann Whitney U test was used to compare polyfunctionality scores between the vaccine groups. Significant associations are shown as ****p < 0.0001; ***p < 0.001; ns,not significant.